Chapter 8. Nematode infections

■ Nematodes, commonly referred to as roundworms, eelworms, or threadworms, are

translucent, non-segmented animals ranging between 0.1mm and several meters in

length

< Diversity of nematode morphology and lifestyle>

■ Many nematode species are associated with insects and the types of relationship

are...

• Facultative parasitism

; some nematodes are able to infect insects as well as having the ability to

complete their life cycle as free-living

• Obligate parasitism (=Pathogens)

; cannot complete life cycle without living insect (usually in the hemocoel)

< Plant parasitic and entomopathogenic nematodes >

http://www.youtube.com/watch?v=D7r1S6-op8E&feature=related

■ Insects serve as vectors or intermediate hosts for a number of nematode

parasites of vertebrates

• mosquitoes transmit Dirofilaria immitis, causal agent of the dog heartworm

< Heartworm infected dog heart >

• black flies transmit Onchocerca volvulus, causal agent of human river blindness

(=onchocerciasis)

< Human onchocerciasis (=river blindness) and causative agent, Onchocerca

volvulus filarial nematode, and black fly vector >

■ Life cycles

• egg

• juvenile (usually have 4 molts J1, J2, J3, J4)

⇨ some insect nematodes have environmentally resistance stage called "dauer

juvenile" (3rd stage juvenile) which ensheathed in the second-stage cuticle

• adult

■ Mode of infection

• insect-parasitic nematodes parasitize their hosts by directly penetrating thru the

cuticle into the hemocoel or by entering thru natural openings (spiracles, mouth,

anus)

• nematodes eggs can be ingested by susceptible hosts and hatched juveniles

penetrate the midgut and enter into the hemocoel

8-1. Mermithid nematodes (Nematoda: Mermithidae)

• infect the early stage of insect hosts (mainly aquatic insects)

• directly penetrate the host's integument or enter via natural openings

(such as spiracle or anus)

• do not reach adulthood in the host's hemocoel

➪ have parasitic (juveniles) and free-living stages (adult)

➪ they emerge from the host as postparasitic juveniles

< Generalized life cycle of mermithid nematodes and emerging postparasitic

juveniles from mosquito larva >

• mermithid nematodes grow until they occupy most of the larval hemocoel,

obtaining all of their nutritional requirements within the host to complete the

parasitic stage of their life cycle

• then the nematode may exit thru the host cuticle, resulting in host death, or

remain in the host thru pupation and into the adult fly

• mermithid infections carrying thru to the adult fly frequently result in structural

and behavioral host feminization

Mermithids emerging

from mosquito larvae

➪ feminization can either be complete or partial (=intersex)

➪ feminized adult black flies exhibit characteristic uninfected gravid female

oviposition behaviors by travelling to upstream oviposition sites and depositing

their mermithids into the water

8-2. Entomopathogenic nematodes (Steinernematidae, Heterorhabditidae)

• obligate pathogens (but recently known that some are facultative scavenger as a

alternative survival strategy)

• recovered from soils throughout the world and their distribution may be primarily

limited by the availability of susceptible hosts

• they can kill the hosts within 1-4 days (usually within 2 days)

• have mutualistic symbiotic bacteria in their guts

- Steinernematid nematodes - Xenorhabdus bacteria

- Heterorhabditid nematodes - Photorhabdus bacteria

• the only stage that survives outside of a host is the non-feeding, non-developing

infective juvenile (IJ) that contain symbiotic bacteria

• in Steinernema species, the symbiotic bacteria are contained inside a

specialized intestinal vesicle (=receptacle), whereas the bacteria of

Heterorhabditis colonize the entire intestine (anterior to mid intestine)

< Location of symbiotic bacteria in intestines of their respective nematode hosts

(A, Steinernema; B, Heterorhabditis) >

• the bacteria usually persist in a quiescent state within the nematode

; some bacteria may grow inside the nematodes

< Xenorhabdus bovienii in Steinernema oregonense >

< Xenorhabdus hominickii in Steinernema monticolum >

1) EPNs and symbiotic bacteria

• nematodes provide symbiotic bacteria with

- shelter

- nutrition

- transportation

• symbiotic bacteria provide nematode with

- toxins to kill the insect host rapidly

➪ some Steinernema spp. can kill the insect host without help of symbiotic

bacteria, but it is less effective

- antimicrobial components (e.g., antibiotics) to protect the cadaver from

contamination by opportunistic organisms including insect scavengers as

well as other common gut inhabiting bacteria including Salmonella,

Pasteurella, and Xanthomonas spp.

- food (bacterivore)

※ All Xenorhabdus and Photorhabdus spp. are symbionts of EPNs only except

Photorhabdus asymbiotica which is isolated from human wounds in the

absence of nematode vectors

< Skin abscess caused by Photorhabdus asymbiotica >

2) Generalized life cycle of EPNs

• the IJs actively search (cruiser) or sit-and-wait (ambusher) for a suitable insect

host and gain entry into the hemocoel by invading through natural openings

(mouth, spiracles, and anus) or, in Heterorhabditis, enter the hemocoel directly

through the insect’s integument

• upon entry into the host, the IJs resume developmental cycle (=recovery) and

liberate their bacterial symbionts

➪ Xenorhabdus are released from the nematodes (Steinernema spp.) by

defecation whereas Photorhabdus exit through the mouth (regurgitation) of

Heterorhabditis spp.

• the bacterial symbionts multiply in the hemolymph and the guts of the host,

producing toxins and exoenzymes that result in septicemia and bioconversion

(=metabolize) of the insect cadaver suitable for nematode feeding

• near the end of bacterial proliferation, the symbionts produce a variety of

antimicrobial compounds that protect the cadaver from colonization by other

organisms

➪ include antibiotics that are active against other bacteria, fungi, and yeasts

and bacteriocins (e.g., xenorhabdicin and lumicins) that are active against

bacteria closely related to Photorhabdus

• the developing nematodes feed on the bacteria and bioconverted host tissues

and reproduce in the cadaver for 1 ~ 3 generations

• once the food resources in the cadaver are exhausted, the nematodes develop

a new generation of IJs that recruit few bacterial cells and emerge from the

host cadaver in search of a new susceptible host

3) Endotokia matricida

• inside + birth + mother + kill

➪ juveniles hatch inside the uterus and feeding on the mother's tissues

causing her death

< Endotokia matricida of Heterorhabditis bacteriophora >

< Endotokia matricida of Steinernema carpocapsae >

• endotokia matricida usually observed in the starved females with not

enough energy to push their eggs thru the reproductive opening

• its function is to provide optimal conditions for offspring development when

the environmental conditions are becoming detrimental

➪ the intra-uterine juvenile development accompanied by the consumption

of the maternal tissues and body content is a favorable and highly

efficient way to raise well-fed IJ, particularly under conditions of nutritive

depletion

4) Steinernematid nematodes (Rhabditida: Steinernematidae)

• both male and female nematodes must enter a susceptible host for

reproduction (sexual)

➪ exception: Steinernema hermaphroditum - has hermaphrodite (as in

Heterorhabditis) in the first generation

• IJs enter a susceptible host via natural openings (do not directly penetrate the

cuticle)

• each species of Steinernema has only single species of bacterial symbiont

➪ Steinernema carpocapsae - Xenorhabdus nematophila

➪ Steinernema monticolum - Xenorhabdus hominickii

• single Xenorhabdus sp. can colonize multiple nematode species

➪ Xenorhabdus bovienii can colonize 8 Steinernema spp.

• signs

- brown to dark brown

- getting flaccid (for some cases, cadavers getting hardened)

- cadaver do not putrefied (decay, rot) because of antibiotics

- characteristic smells (depend on Xenorhabdus spp.)

< Galleria mellonella larvae infected with Steinernema sp. >

< Flaccid cadaver caused by Steinernema jollietti >

EPN from bait insect cadaver

• Steinernema nematodes can be cultured in an artificial medium

< Lipid agar medium for Steinernema nematode in vitro culture >

⇒ this doesn't mean that they are not obligate parasites

5) Heterorhabditid nematodes (Rhabditida: Heterorhabditidae)

• all infective juveniles become hermaphrodites

➪ only single individual is required to infect a new host for successful

reproduction

• IJs infect a susceptible host by direct penetration of the cuticle

< Typical discoloration and bloating of Heterorhabditis infected larvae >

• signs

- cadaver turn to reddish dark brown

- no typical smells

- cadaver usually bloated

6) EPNs and biological control

• many EPNs have been used as biological control agent for various insect

pests including lepidopteran larvae, mole crickets, and fly larvae

• few EPNs have a broad host range (e.g., Coleoptera, Lepidoptera, Diptera

larvae)

➪ Steinernema carpocapsae, Steinernema riobrave, Heterorhabditis

bacteriophora

• some EPNs have a restricted host range

➪ Steinernema scapterisci is specialized to cricket and mole crickets

➪ Steinernema feltiae is specialized to fly larvae

< Fungus gnat larvae infected by S. feltiae >

➪ Steinernema kushidai and Steinernema scarabaei are restricted to scarab

larvae

; other species such as Steinernema glaseri and Heterorhabditis

bacteriophora also usually infect scarab larvae in natural environment

• Host finding strategies

; EPNs are positively attracted to carbon dioxide

- ambusher(sit and wait)

- cruiser

• Effect of temperature

- 10∼15℃: getting sluggish

- 30∼40℃: inactive

- 20∼30℃: optimum for most species

• Application of EPNs in the field

- when EPNs applied onto the soil surface by using conventional straying

equipment, losses can reach 50% within hours of application due primarily

to UV radiation and desiccation

- generally only 1% of the original inoculum survives after 1∼6 weeks

- it is important that EPN should only be applied when susceptible stages of

the target pest are present